The goal of this project is to generate recombinant polyclonal antibody libraries (PCALs) that could be used, alone or in part, for prophylactic or therapeutic treatment of tularemia, as well as for diagnosis or environmental detection. Francisella tularensis, the etiologic agent of tularemia, is a facultative intracellular bacterium that causes a debilitating, sometimes fatal disease. Despite successful treatment with antibiotics, the morbidity associated with infection and the threat of engineered multiple antibiotic-resistant strains for bioterrorism, suggest the need for additional strategies to combat tularemia. We have developed the PCAL technology to combine the advantages of serum-derived polyclonal antibodies and hybridoma-derived monoclonal antibodies. Hence, PCALs are standardized mixtures of polyclonal antibodies for which the genes are available and therefore the antibodies can be amplified and modified as desired. Antibody gene repertoires are harvested from B cell-containing tissues by RT-PCR and cloned to produce Fab phage display libraries that can be positively and negatively selected for desired binding specificities. The selected libraries are then converted, by mass gene transfer, into mammalian expression vector libraries encoding full-length antibodies in which the same repertoire of antibody variable region genes can be connected to constant region genes of any isotype or species. PCALs are then produced by transfection into mammalian cells. The specific aims for the current proposal are: 1) Optimize the antibody response in mice to virulent strains of F. tularensis; 2) Produce F. tularensis-specific PCALs that are all-mouse, all-human, or chimeric, from mice optimally immunized with F. tularensis; 3) Determine the in vitro efficacy of the PCALs produced in Aim 2 in complement-mediated killing and opsonophagocytosis (with or without complement) assays and in blocking infection of macrophages in culture; 4) Determine the in vivo prophylactic and therapeutic efficacy of the PCALs produced in Aim 2 against different strains of F. tularensis in mouse models of inhalational infection. The PCALs generated in this proposal will be available for industrial scale-up for prophylactic/therapeutic use as well as for diagnostic and environmental detection devices.